Relaxin-3 null mutation mice display a circadian hypoactivity phenotype.

Characterizing the neurocircuits and neurotransmitters that underlie arousal and circadian sleep/wake patterns is an important goal of neuroscience research, with potential implications for understanding human mental illnesses, such as major depression. Recent anatomical and functional studies suggest that relaxin-3 neurons and their ascending projections contribute to these functions via actions on key cortical, limbic and hypothalamic circuits. This study reports the behavioral phenotype of C57BL/6J backcrossed relaxin-3 knockout (KO) mice. Cohorts of adult, male and female relaxin-3 KO and wild-type (WT) littermate mice were subjected to a battery of behavioral tests to assess sensorimotor function and complex behavior. No overt deficits were detected in motor-coordination, spatial memory, sensorimotor gating, anxiety-like behavior or locomotor behavior in novel environments; and no marked genotype differences were observed in response to a chronic stress protocol. Notably however, compared to WT mice, relaxin-3 KO mice displayed robust hypoactivity during the dark/active phase when provided with free home-cage access to voluntary running wheels. This circadian hypoactivity was reflected by reduced time spent and distance traveled on running wheels, coupled with an increase in the time spent immobile, possibly reflecting increased sleeping. Overall, these studies support a role for relaxin-3 signaling in the control of arousal and sleep/wakefulness, and identify the relaxin-3 KO mouse as a useful model to study this role further.

Relaxin-3 in GABA projection neurons of nucleus incertus suggests widespread influence on forebrain circuits via G-protein-coupled receptor-135 in the rat.

Relaxin-3 (RLX3) is a newly identified member of the relaxin/insulin peptide family that is highly conserved across a range of species from fish to mammals and is highly expressed in rat, mouse and human brain. Extensive pharmacological studies have demonstrated that RLX3 is a high affinity, selective ligand for G-protein-coupled receptor-135 (GPCR135, now classified as relaxin family peptide-3 receptor; RXFP3). In ongoing studies to understand the physiological functions of RLX3, the distribution of RLX3-containing neuronal elements in rat brain was determined by immunohistochemistry, using an affinity-purified polyclonal antiserum raised against a conserved segment of the RLX3 C-peptide (AS-R3(85-101)). Consistent with the distribution of RLX3 mRNA, neurons containing RLX3-like immunoreactivity (LI) were observed in the pontine nucleus incertus and the majority of these cells, which are known to express corticotropin-releasing factor receptor-1, were shown to express glutamic acid decarboxylase-65-immunoreactivity, suggesting a GABA phenotype. Nerve fibers and terminals containing RLX3-LI were observed adjacent to cells in the nucleus incertus and in various forebrain regions known to receive afferents from the nucleus incertus, including cortex, septum, hippocampus, thalamus, hypothalamus and midbrain. Regions that contained highest densities of RLX3-positive fibers included the medial septum, lateral preoptic area, lateral hypothalamus/medial forebrain bundle and ventral hippocampus; and additional fibers were observed in olfactory bulb and olfactory and frontal/cingulate cortices, bed nucleus of the stria terminalis, dorsal endopiriform, intergeniculate, and supramammillary nuclei, and the periaqueductal gray and dorsal raphe. The RLX3-positive network overlapped the regional distribution of GPCR135 mRNA and specific binding sites for an [125I]-GPCR135-selective, chimeric peptide. These anatomical findings further support the proposition that RLX3 is the endogenous ligand for GPCR135 in rat brain and provide evidence for broad modulatory activity of RLX3 in behavioral activation relating to autonomic and neuroendocrine control of metabolism and reproduction and higher-order processes such as stress and cognition.

Clinical benefits of continuous leukocyte filtration during cardiopulmonary bypass in patients undergoing valvular repair or replacement.

Valve operations in the form of repair or replacement make up a significant population of patients undergoing surgical procedures in the USA annually with the use of cardiopulmonary bypass. These patients experience a wide range of complications that are considered to be mediated by activation of complement and leukocytes. The extracorporeal perfusion circuit consists of multiple synthetic artificial surfaces. The biocompatibility of the blood contact surfaces is a variable that predisposes patients to an increased risk of complement mediation and activation. This can result in an inflammatory process, causing leukocytes to proliferate and sequester in the major organ systems. The purpose of this study was to determine whether filtration of activated leukocytes improved clinical outcomes following surgical intervention for valve repair or replacement. In this paper, we report a retrospective matched cohort study of 700 patients who underwent valve procedures from June 1999 to December 2002. The control group (CG) consisted of patients who had a conventional arterial line filter. In the study group (SG), patients had a conventional arterial line filter and a leukocyte arterial line filter (Pall Medical, NY). In the SG, blood diverted to the cardioplegia system was also leukocyte depleted to enhance myocardial preservation by adapting this device to the outflow port on the filter. Patient characteristics were similar for the SG and the CG, including 228 males and 122 females, mean age (62.4 versus 64.2 years), cardiopulmonary bypass time (127+/-64 versus 116+/-53 min), and aortic crossclamp time (84+/-23 versus 81+/-23 min). Our results demonstrate that the SG achieved statistically significant reduction in the time to extubation (p =0.03) and the number of patients with prolonged intubation in excess of 24 hours (p <0.04), in addition to improved postoperative oxygenation (p=0.01), and decreased length of hospital stay (p =0.03). We believe that leukocyte filters are clinically beneficial, as demonstrated by the results presented in this study.

Intraoperative modality of treatment for peritoneal carcinomatosis: use of hyperthermic interperitoneal chemoperfusion.

The use of hyperthermia as an adjunct to chemotherapy in the treatment of peritoneal carcinomatosis is a promising technique for patients who otherwise have a poor prognosis for survival. We, herein, report an overview and description of our technique for the safe conduct of this treatment. Included in these data are a total of 71 patients who underwent an intraoperative treatment with Mitomycin C at temperatures of 41-42 degrees C for a 90- to 120-min time period. The treatment protocol, perfusion system description, technical considerations, and potential complications are also included. The prognosis for intraabdominal carcinomatosis is poor with conventional treatments and modalities. We believe that the use of this technique offers a very positive clinical alternative for patients undergoing treatment for laparoscopic palliation of malignant ascites and/or surgical debulking for intraoperative treatment and prevention of metastasis.

Anatomical distribution of prolactin-releasing peptide and its receptor suggests additional functions in the central nervous system and periphery.

A recently identified neuropeptide with PRL-releasing capabilities binds to and activates a previously known orphan G protein-coupled receptor, GPR10. We initiated a study to define the pharmacology of the peptide/receptor interaction and to identify the distribution of the peptide and its receptor in the central nervous system to elucidate sites of action of the peptide. The PRL-releasing peptide (PrRP) is a C-terminally amidated, 31-amino acid peptide derived from a 98-amino acid precursor. Radioiodinated PrRP-(1-31) binds to its receptor with high affinity (1 nM) and stimulates calcium mobilization in CHOK1 cells stably transfected with the receptor. A series of N-terminal deletions reveals that the PrRP-(12-31) amino acid is equipotent to PrRP-(1-31). Further N-terminal deletions reduce the affinity of the ligand considerably, although PrRP-(25-31) is still able to compete for binding and behaves as an agonist. The arginine residues at position 26 and 30 are critical for binding, as substitution with either lysine or citrulline reduces the affinity substantially. In situ hybridization reveals a distinct tissue distribution for both the peptide and receptor messenger RNAs. The receptor is expressed abundantly in the reticular thalamic nucleus, periventricular hypothalamus, dorsomedial hypothalamus, nucleus of the solitary tract, area postrema, anterior pituitary, and adrenal medulla. The peptide messenger RNA is expressed in the dorsomedial hypothalamus, nucleus of the solitary tract, ventrolateral reticular nucleus, and intestine. This tissue distribution suggests an alternative function of PrRP than its purported hypophysiotropic function, such as a potential role for PrRP in the central feedback control of neuroendocrine and autonomic homeostasis. Further work using selective agonists and antagonists should help define additional physiological roles of this novel mammalian neuropeptide.

Comparison of an agonist, urocortin, and an antagonist, astressin, as radioligands for characterization of corticotropin-releasing factor receptors.

The characteristics of a high-affinity antagonist radioligand are compared with those a high-affinity agonist in binding to the cloned corticotropin-releasing factor receptor type 1 (CRF-R1) and type 2 (CRF-R2) and to the native receptors that exist in rat cerebellum and brain stem. The relative potencies of CRF antagonists and agonists to the two types of cloned CRF receptors overexpressed stably in Chinese hamster ovary cells are determined using the antagonist radioligand 125I- [DTyr1]astressin (Ast*), and the agonist radioligand, 125I -[Tyr0]rat urocortin (Ucn*). The inhibitory binding constants (Ki) of astressin and urocortin are 1 to 2 nM for all receptors and are independent of which radioligand is employed. Astressin binds with high affinity to the native cerebellar/brain stem receptor and relative potencies of selected CRF analogs determined with Ast* on the native receptor are similar to those obtained for the cloned CRF-R1. The specific binding of Ast* to endogenous brain receptors is greater than that of Ucn*, resulting in more sites being detected by the antagonist than by the agonist. In contrast to another CRF agonist, the binding of Ucn* to the cloned receptors is relatively insensitive to guanyl nucleotides at both 20 degreesC and 37 degreesC; however, its binding to the native receptor is displaced by guanyl nucleotides at 37 degreesC and, to a lesser degree, at 20 degreesC. As expected, the binding of the antagonist Ast* is not affected by guanyl nucleotides. Because it is a high-affinity, specific CRF antagonist, astressin is eminently suitable as a ligand for detection and characterization of both endogenous and cloned CRF receptors.

Sickle cell disease and aortic valve replacement: use of cardiopulmonary bypass, partial exchange transfusion, platelet sequestration, and continuous hemofiltration.

Sickle cell disease in patients undergoing open heart procedures presents a multitude of challenges to the medical staff. With improved techniques of cardiopulmonary bypass, surgery, and anesthesia for treating patients with sickle cell disease, perfusionists will likely encounter patients with this genetic disorder on a more frequent basis. A 40-year-old black woman was admitted to our institution with recurrent Staphylococcus epidermidis and sepsis. She underwent transesophageal echocardiography and cardiac catheterization and was subsequently diagnosed with severe aortic insufficiency. The aortic valve was replaced. Herein, we report our experience in the preoperative, perioperative, and postoperative management of this patient. We present a concise update on the current literature and techniques used by others in similar cases, and we provide a brief section on future considerations to assist fellow practitioners in recognizing this disease and meeting the accompanying challenges.

Cloning and characterization of human urocortin.

Urocortin, a new member of the CRF peptide family which also includes urotensin I and sauvagine, was recently cloned from the rat midbrain. The synthetic replicate of urocortin was found to bind with high affinity to type 1 and type 2 CRF receptors and, based upon its anatomic localization within the brain, was proposed to be a natural ligand for the type 2 CRF receptors. Using a genomic library, we have cloned the human counterpart of rat urocortin and localized it to human chromosome 2. Human and rat urocortin share 95% identity within the mature peptide region. Synthetic human urocortin binds with high affinity to CRF receptor types 1, 2 alpha, and 2 beta, stimulates cAMP accumulation from cells stably transfected with these receptors, and acts in vitro to release ACTH from dispersed rat anterior pituitary cells. In addition, the CRF-binding protein binds human urocortin with high affinity and can prevent urocortin-stimulated ACTH secretion in vitro. The inhibitory effect of the CRF-binding protein on human urocortin can be blocked by biologically inactive CRF fragments, such as CRF(9-33).

Ligand requirements of the human corticotropin-releasing factor-binding protein.

CRF-binding protein (CRF-BP), identified as a 37-kilodalton human serum protein, binds human (h) CRF (Kd = 0.17 +/- 0.01 nM) and blocks hCRF's ability to stimulate ACTH release by pituitary cells in vitro. The present study examines ligand requirements of CRF-BP by testing the affinity of recombinant CRF-BP for synthetic analogs of CRF and peptides in the CRF family. The relative affinities of various fragments of hCRF or related peptides for CRF-BP indicate that residues 9-28 are crucial for ligand binding. CRF-BP binds human/rat CRF and urotensin-I with high affinity, sauvagine with moderate affinity, and ovine (o) CRF with low affinity. The marked difference in the affinity of CRF-BP for oCRF (Ki = 1100 +/- 97 nM) compared to hCRF (Ki = 0.17 +/- 0.01 nM), when considered with the importance of the central domain, suggests that amino acids 22, 23, and/or 25 are critical for binding. Altering oCRF residues 22, 23, or 25 individually or collectively to match those of hCRF increases the affinity of CRF-BP for these ligands; [Ala22, Arg23, Glu25]oCRF, in which all three of these central amino acids are substituted by their hCRF counterparts, binds CRF-BP with an affinity equal to that of hCRF. CRF-BP has differential affinities for CRF receptor antagonists, binding alpha-helical CRF-(9-41) with high affinity and [D-Phe12, Nle21,38]hCRF-(12-41) with low affinity. Thus, the structural requirements for binding to CRF-BP can clearly be distinguished from those for CRF receptor recognition of both agonists and antagonists. Peptides such as hCRF-(9-33), with low biological activity but which retain high affinity for the binding protein, can competitively override the effects of CRF-BP to block CRF-induced ACTH secretion, raising the possibility that whereas endogenous CRF-BP serves to limit the distribution or duration of action of CRF, specific pharmacological inhibitors of the ligand-binding protein interaction might be used to therapeutically elevate free CRF levels.

Central activin administration modulates corticotropin-releasing hormone and adrenocorticotropin secretion.

A broad and diffuse neuronal network conveys information reflecting the state of the internal and external environment to the neurosecretory hypothalamus. Recently, we identified an inhibin-beta A- (I beta A) immunoreactive terminal field within the CRF-rich portion of the dorsomedial paraventricular nucleus which originates from a cell group in the commissural portion of the nucleus of the solitary tract (NTS). The NTS receives baroreceptor input, somatosensory input via the spinosolitary tract, and sensory information from the oral, thoracic, and abdominal cavities and, thus, is positioned to serve as a primary relay for visceral sensory inputs to neurons critical to the function of the hypothalamic-pituitary-adrenal (HPA) axis. Although these NTS cells contain multiple putative transmitters, we present evidence that activin, an inhibin-beta A dimer, plays a modulatory role in HPA axis function via facilitation of CRF release. First, intraventricular injection of activin-A (0-3 nmol), but not the related inhibin heterodimer, evoked dose-related 1.7- to 2.8-fold elevations of circulating ACTH levels in male rats. Second, analysis of hypophysial-portal plasma after bilateral paraventricular nucleus microinfusion of activin-A revealed a dose-related facilitation of CRF secretion up to 4-fold above preinjection levels which was unaccompanied by changes in arginine vasopressin levels. Finally, activin-A also enhanced CRF secretion from neonatal hypothalamic cells in primary culture with an EC50 dose of approximately 0.25 nM. Overall, these observations provide evidence of both an anatomical and a pharmacological substrate for activin-mediated central modulation of HPA axis function.

Facilitatory role of neuropeptide Y on the onset of puberty: effect of immunoneutralization of neuropeptide Y on the release of luteinizing hormone and luteinizing-hormone-releasing hormone.

To examine the role of neuropeptide Y (NPY) in the first luteinizing hormone (LH) surge of puberty, the effect of passive immunoneutralization of NPY with antiserum against NPY (anti-NPY) injected centrally (third ventricle) or peripherally (jugular vein) was studied in pubertal female rats on the day of first proestrus. Both peripheral and central anti-NPY administration reduced the magnitude of the LH surge during the afternoon of first proestrus; however, the central route of administration appeared to be most effective. Centrally administered anti-NPY also reduced the magnitude of proestrous LH-releasing hormone (LHRH) release into pituitary portal blood in these rats. These results suggest that endogenous NPY plays a facilitatory role in the generation of the LHRH surge necessary for preovulatory gonadotropin release and puberty.

Glucocorticoid feedback inhibition of adrenocorticotropic hormone secretagogue release. Relationship to corticosteroid receptor occupancy in various limbic sites.

Feedback inhibition of the adrenocortical axis by circulating glucocorticoids occurs at the pituitary and CNS sites. In the CNS, both hypothalamic and suprahypothalamic sites have been implicated as mediators of glucocorticoid feedback activity. In the present experiments, we have attempted to identify specific CNS regions mediating the feedback and to characterize which hypothalamic adrenocorticotropic hormone secretagogues are under glucocorticoid inhibitory control. Adrenalectomized rats were presented with a delayed feedback signal in the form of systemic infusion with corticosterone or dexamethasone. Hypophysialportal concentrations of corticotropin-releasing factor (CRF), arginine vasopressin (AVP), and oxytocin (OT) were determined before and during a hypotensive stressor in the face of varying levels of feedback. The rats were then killed, and the extent of total, type I, and type II corticosteroid receptor occupancy in hippocampus, hypothalamus, and amygdala was determined. The following observations were made: (1) increased hippocampal corticosteroid receptor occupancy was associated with suppressed adrenocorticotropic hormone secretagogue concentrations; (2) the major, significant predictor of initial (prehypotensive) concentrations of CRF, AVP, and OT was the extent of occupancy of hippocampal type II receptors, often in combination with occupancy of hippocampal type I or hypothalamic receptors; (3) secretion of CRF induced by hypotension was best predicted by hippocampal type I and type II receptor occupancy (stress-induced OT secretion was best predicted by hippocampal type II and hypothalamic receptor occupancy), and (4) the 'shape' of the hippocampal type II receptor occupancy versus initial AVP concentration curve suggested a nonlinear, threshold type of relationship, implying tight hippocampal regulation of AVP secretion.

Elevation of hypophysial portal concentrations of adrenocorticotropin secretagogues after fornix transection.

Glucocorticoid feedback inhibition at the level of the brain is extremely complex, involving feedback at both hypothalamic and suprahypothalamic levels. The hippocampus has been implicated as a suprahypothalamic mediator of such feedback, based on numerous lesion, stimulation, and steroid implantation studies. These reports, however, predated the isolation and characterization of CRF and recognition of the multifactorial control of ACTH release. Thus, it is not clear which hypothalamic ACTH secretagogues are under inhibitory control of the hippocampus. To answer this, we measured hypophysialportal concentrations of CRF, arginine vasopressin, and oxytocin in rats with fornix transections, which disrupt hippocampal communication with the hypothalamus. Hypophysial-portal blood was collected in rats exposed to either low or high circulating corticosterone concentrations in the presence or absence of the coincident stressor of hypotension. We observed that fornix transection produced hypersecretion of all three secretagogues. However, the pattern of hypersecretion differed for each as follows: 1) fornix transection did not affect either initial CRF secretion or the magnitude of the stress response, but made rats resistant to a high feedback signal during stress; 2) fornix transection led to initial arginine vasopressin hypersecretion, which remained sensitive to a high feedback signal; and 3) fornix transection led to initial oxytocin hypersecretion as well as resistance to a high corticosterone feedback signal during hypotension.

Neuropeptide Y (NPY): a possible role in the initiation of puberty.

NPY modulates the rat hypothalamic-pituitary-gonadal axis at both the adenohypophysial and central levels. Previously published studies have consistently shown elevations of GnRH content in the preoptic area (POA) and hypothalamus starting with the appearance of GnRH immunoreactivity around fetal day 12-14 until stabilization around the time of puberty. In the present studies, irNPY content of male and female rat hypothalami and POA was examined during days 0 to 36 of postnatal development. Hypothalamic irNPY content rose steadily from 4.54 +/- 0.19 ng/fragment (males) and 2.72 +/- 0.55 ng/fragment (females) at birth to 34.14 +/- 3.94 ng/fragment (males) and 46.79 +/- 6.16 ng/fragment (females) at day 36, corresponding approximately to the time of vaginal opening. A similar elevation of irNPY content was observed in the POA. At day 0, POA content was 1.91 +/- 0.18 ng/fragment (males) and 2.02 +/- 0.25 ng/fragment (females) and progressively increased to 42.26 +/- 3.94 ng/fragment and 41.33 +/- 3.72 ng/fragment by postnatal day 36. In subsequent investigations, hypophysial-portal and peripheral plasma irNPY was determined around the time of vaginal opening, revealing a surge in portal levels of irNPY which preceded the prepubertal LH surge. The progressive postnatal increase in hypothalamic and POA irNPY content culminating in a prepubertal surge of irNPY secretion into the hypophysial-portal circulation suggests involvement of this neuropeptide in reproductive development and the onset of puberty.

Regulation of corticotropin-releasing factor secretion in vitro by glucose.

The neurosecretory responses of the isolated rat hypothalamus were assessed in vitro. Rat hypothalamic blocks were incubated for 30 min in a N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered salt solution with 5.5 mM glucose (base-line collection period). The blocks were transferred to fresh buffer with a new concentration of glucose with or without various additions (test period); corticotropin-releasing factor (CRF) and other hormones in the media were determined by radioimmunoassay. CRF secretion was maximally increased to approximately 200% of base line at glucose concentrations less than 4 mM and decreased to 65% of base line at higher glucose concentrations. The increase in CRF secretion at low glucose (0.55 or 1.38 mM) was Ca2+ dependent and completely reversible. Hexamethonium, cyproheptadine, and atropine partially blocked the CRF response to 0.55 mM glucose. Glucose concentrations from 0 to 11 mM had no effect on the CRF response to 47.5 mM KCl. The inhibitory effects of high glucose were completely reversed by the addition of 2-deoxy-D-glucose (3-49 mM). Glucose levels did not alter secretion of either gonadotropin-releasing hormone or arginine vasopressin from hypothalamic blocks. The results suggest that the isolated rat hypothalamus is extremely sensitive to the level of glucose and that CRF is rapidly and reversibly secreted in response to slight reductions in glucose concentrations. These concentrations are consistent with those observed during moderate to severe hypoglycemia in vivo. The rise in glucocorticoids observed in vivo during hypoglycemia may result at least in part from the ability of the hypothalamus to directly sense glucose levels and promote secretion of CRF.

Evidence that neuropeptide Y (NPY) released into the hypophysial-portal circulation participates in priming gonadotropes to the effects of gonadotropin releasing hormone (GnRH).

NPY exhibits modulatory activity at both the adenohypophysial and the central levels of the rat hypothalamic-pituitary-gonadal axis. In the present studies, the secretion and physiological activity of endogenous irNPY was examined. We have shown that: (i) irNPY is secreted into the hypophysial-portal circulation, (ii) hypophysial-portal concentration profiles of irNPY and irGnRH are parallel throughout the rat estrous cycle, and (iii) removal of endogenous NPY via immunoneutralization inhibits the steroid-induced LH surge in ovariectomized rats. From these observations, we speculate that NPY secreted into the hypophysial-portal circulation participates in priming of gonadotropes to the actions of GnRH on the afternoon of the preovulatory surge.

Analysis of the role of angiotensin II in mediation of adrenocorticotropin secretion.

It has been suggested that ACTH secretion in response to selected stimuli may be modulated by angiotensin II (AII) via direct action at the level of the corticotrope or through central actions to facilitate CRF secretion into the hypophysial-portal circulation. These hypotheses were evaluated in the present series of experiments. Our failure to observe a significant portal to peripheral plasma immunoreactive (ir) AII gradient suggested that AII does not act as a physiologically significant ACTH secretagogue at the level of the corticotrope. Central administration of synthetic AII evoked a modest dose-related stimulation of hypothalamic irCRF secretion into the portal circulation, which was reversed by the AII receptor antagonist saralasin. Activation of a known AII-positive neuronal pathway projecting from the subfornical organ (SFO) to the hypothalamic paraventricular nuclei resulted in an elevation of hypophysial-portal plasma irCRF levels and increased circulating ACTH. Pretreatment with saralasin prevented SFO stimulation-induced irCRF secretion. These observations suggest that central AII-containing pathways may participate in mediation of ACTH secretion in response to hypovolemia or hyperosmolality by facilitating irCRF secretion. Involvement of the SFO, a circumventricular organ, in this circuit is intriguing as it provides a means of monitoring peripheral irAII concentration and then converting this humoral signal into a neural signal distributed to regions regulating drinking behavior, neurohypophysial AVP secretion, and activation of the hypothalamic-pituitary-adrenal axis.

Central administration of corticotropin-releasing factor modulates oxytocin secretion in the rat.

Recently, it has been reported that oxytocin (OT), classically known for its function during parturition and lactation, is secreted in response to stressful stimuli in male rats. In these and in the present report it was found that swimming stress, restraint stress, ether stress, and footshock stress elevate OT secretion without affecting arginine-vasopressin (AVP) secretion. In the present studies, we investigated the possible modulation of OT secretion by CRF which is known to be released during stress. Male and female rats received intraventricular (icv) injections of 0.75 nmol (5 micrograms) rat CRF and were killed 5 min after the treatment. CRF significantly elevated OT secretion in male and female rats 3.4- and 4-fold, respectively. Plasma AVP levels were not affected by the treatment. The effect of CRF on OT release was structure specific since rat CRF, ovine CRF, and sauvagine were equipotent releasers of OT while an inactive analog to CRF, ovine CRF did not change plasma OT levels. In another set of experiments rats were pretreated with either CRF-antiserum (0.5 ml iv) or dexamethasone (20 micrograms/rat ip) and then injected with icv CRF. Both CRF-antiserum and dexamethasone blocked the rise in ACTH release after icv CRF completely but did not influence the OT response. This suggests CRF may be acting centrally but not at the level of the neurohypophysis to change OT secretion. Since parvocellular but not magnocellular neurons of the paraventricular nucleus have been demonstrated to be steroid sensitive in immunohistochemical studies, we suggest CRF may act directly or indirectly upon magnocellular neurons to increase OT release. Intravenous administration of 0.75 nmol CRF increased both OT and AVP levels in peripheral blood. The magnitude of this increase was similar (2- to 4-fold stimulation) to responses after icv administration of CRF. Intravenous administration of CRF results in hypotension and may therefore cause a baroreceptor mediated release of AVP and OT. From the above evidence we conclude: physical and mental stresses which do not result in changes in blood volume or osmolality evoke an increase in OT secretion while AVP secretion remains unchanged; CRF administered icv mimics OT responses observed after ether stress or footshock stress; CRF may play a role in regulating stress-induced OT secretion in the rat.